The present invention relates to oligonucleotides, kits, microarrays, and methods for detection of cardiovascular disease, in particular, left ventricular heart failure.
The World Health Organization has determined that cardiovascular disease (CVD) is the leading cause of death throughout the world and the leading cause of lost years of healthy life in Europe. In the U.S., CVD caused 41 per cent of all deaths in 1998 and is second only to all cancers combined in years of potential life lost. The annual number of deaths from CVD in the U.S. increased substantially between 1900 and 1970, peaking in 1963. Since 1963 the U.S. CVD death rate has changed from an increasing to a decreasing trend, and by 1995 the CVD death rate was similar to that in 1936.
The only exception to the CVD mortality decline in the U.S. is congestive heart failure (CHF). CHF is often the end stage of cardiac disease, and half of the patients diagnosed with CMF die within five years. Sudden death is common among CHF patients, occurring six to nine times more frequently than in the general population. Between 1968 and 1998, the number of deaths from CHF in the U.S. increased from approximately 10,000 to almost 50,000. Approximately five million Americans are currently living with heart failure, and 550,000 new cases are diagnosed in the U.S. each year. CHF occurs slightly more frequently among men than among women, and is twice as common among African Americans compared to Caucasian Americans, with mortality also doubled for African Americans. The number of CHF cases is expected to increase as the population ages, and as cardiac patients are able to survive and live longer with their disease. CMF has therefore become a major medical problem in the U.S.
As a symptom of underlying heart disease, heart failure is closely associated with the major risk factors of CVD: smoking, high cholesterol, hypertension, diabetes, abnormal blood sugar levels, and obesity, with hypertension and diabetes being particularly important. CMF is about twice as common for persons with hypertension as compared with normotensive individuals, and the degree of risk for heart failure appears to be directly related to the severity of high blood pressure. Persons with diabetes have a two- to eightfold greater risk for heart failure than those without diabetes, and women with diabetes have a greater risk of heart failure than men with diabetes. The presence of coronary disease is one of the greatest risks for heart failure. Muscle damage and scarring caused by heart attacks create a fivefold increase in risk of developing CHF.
Heart failure occurs when one or more of the chambers of the heart cannot keep pace with the amount of blood flowing through an individual""s body. Heart failure can involve either or both sides of the heart, but in general, the left side of the heart is affected first. Each side of the heart comprises an atrium (the upper chamber) which receives blood into the heart and a ventricle (the lower chamber) which pumps blood to the body. The left ventricle supplies most of the heart""s pumping power and is larger than other heart chambers, pumping oxygen-rich blood from the left atrium to the rest of the body. When the left ventricle loses its ability to contract, the heart is unable to pump with sufficient force to push enough blood into circulation. This condition is known as systolic failure. When the left ventricle loses its ability to relax, usually because of stiffness from thickening (hypertrophy), the heart cannot fill properly with blood during the resting period between each heartbeat. This condition is known as diastolic failure. In either condition, blood entering the left atrium from the lungs may back up or regurgitate, causing fluid to leak into the lungs (pulmonary edema), and blood flow throughout the body slows down, causing fluid to build up in tissues (edema). The presence of excess fluid due to heart failure is termed congestive heart failure.
The symptoms of heart failure include shortness of breath (dyspnea), fatigue, fluid accumulation, and persistent coughing that produces mucus or pink, blood-tinged sputum. Heart failure develops slowly, and the heart compensates as its pumping capacity decreases. For example, the heart may enlarge, or muscle fibers within the heart may hypertrophy to allow the heart to contract with more force and pump more blood. The heart also compensates for loss of pumping capacity by contracting more frequently to increase circulation. However, eventually the heart can no longer compensate and symptoms only appear when the condition has progressed in severity.
Physicians currently diagnose heart failure through physical examination, to detect the observable symptoms indicated above. Electrocardiography is performed to determine whether the patient""s heartbeat is abnormal and to identify the cause of the heart failure. In particular, an electrocardiogram with Q-waves and poor R wave progression indicates a previous myocardial infarction, with probable left ventrical systolic dysfunction. An electrocardiogram indicating left ventricular hypertrophy may show an etiology related to hypertension, aortic stenosis, hypertrophic cardiomyopathy or dilated cardiomyopathy, and may indicate either systolic or diastolic left ventricular dysfunction. A chest x-ray may indicate pulmonary edema, pulmonary venous engorgement, or cardiac enlargement. However, heart failure from left ventricular dysfunction may be present even when a patient has a normal chest x-ray. Physicians also perform an ultrasonic examination of the heart to generate an echocardiogram, which allows measurement of systolic and diastolic ventricular contractile function, chamber size, and wall thickness. When Doppler ultrasound is used for echocardiography, valvular stenosis and regurgitation can be detected and quantified.
Lifestyle changes such as weight reduction; moderate exercise; limiting fluid intake, including alcohol intake; and cessation of smoking, are indicated for individuals with chronic CHF. Left ventricular systolic heart failure can be treated using lifestyle changes, angiotensin converting enzyme inhibitors, diuretics, and/or digoxin. The optimal treatment for left ventricular diastolic heart failure has not been determined. Since left ventricular diastolic heart failure is frequently associated with hypertension, antihypertensive therapy is frequently prescribed, and congestion is relieved with diuretics.
A need remains for detection of predisposition to heart failure in individuals who may have the condition but who do not exhibit symptoms, so that lifestyle changes can be initiated early in the progression of disease. In addition, it is important to determine predisposition to heart failure in individuals who have experienced myocardial infarction, so that appropriate treatment modalities may be initiated to limit the effects of heart failure in such individuals.
The present inventors have found that predisposition to left ventricular heart failure may be detected by analysis of certain polymorphisms in the nucleic acid set forth in SEQ ID NO:1. Specifically, the presence of an A nucleotide at position 24801 of SEQ ID NO:1 (the sense strand), or a T nucleotide at the corresponding position of the complement of SEQ ID NO:1, is indicative of a predisposition to left ventricular diastolic heart failure. Furthermore, the presence of a T nucleotide at position 24941 of SEQ ID NO:1, or an A at the corresponding position in the complement of SEQ ID NO:1, is also indicative of predisposition to left ventricular diastolic heart failure. In addition, the presence of an A nucleotide at position 32614 of SEQ ID NO:1 (the sense strand), or a T nucleotide at the corresponding position of the complement of SEQ ID NO:1, is indicative of a predisposition to left ventricular systolic heart failure in individuals who have experienced a myocardial infarction.
Moreover, individuals who possess the SEQ ID NO:1 haplotype characterized by a G nucleotide at position 24801 of SEQ ID NO:1; a C nucleotide at position 24941 of SEQ ID NO:1; a C nucleotide at position 27645 of SEQ ID NO:1; a C nucleotide at position 32163 of SEQ ID NO:1; and a G nucleotide at position 32614 of SEQ ID NO:1 are at lower risk of developing left ventricular diastolic heart failure.
The nucleic acid of SEQ ID NO:1 (nucleotides 1-38653 of GenBank Accession No. AC004923) is the genomic sequence of a gene of unknown function currently described as hUNC93B1 (Kashuba, et al., submitted for publication). The hUNC93B1 gene is located on chromosome 11q13, and the mRNA (GenBank Accession No. AJ271326, SEQ ID NO:50) is expressed at high levels in cardiac tissue (Id.). Without wishing to be bound by any theory, it is believed that the hUNC93B1 gene may encode a twelve transmembrane transporter protein.
In one embodiment, the invention provides a sequence determination oligonucleotide complementary to a polymorphic region within a nucleic acid having a sequence as set forth in SEQ ID NO:1, wherein the region corresponds to a polymorphic site selected from the group consisting of position 24801 of SEQ ID NO:1, position 24941 of SEQ ID NO:1, position 27645 of SEQ ID NO:1, position 32163 of SEQ ID NO:1, and position 32614 of SEQ ID NO:1.
In yet another embodiment, the invention provides a microarray comprising at least one oligonucleotide complementary to a polymorphic region in the nucleic acid set forth in SEQ ID NO:1, wherein the region corresponds to a polymorphic site selected from the group consisting of position 24801 of SEQ ID NO:1, position 24941 of SEQ ID NO:1, position 27645 of SEQ ID NO:1, position 32163 of SEQ ID NO:1, and position 32614 of SEQ ID NO:1.
In another embodiment, the invention provides the oligonucleotide primer pairs useful for amplification of a polymorphic region in the nucleic acid of SEQ ID NO:1 from a biological sample, wherein the region corresponds to a polymorphic site selected 10 from the group consisting of position 24801 of SEQ ID NO:1, position 24941 of SEQ ID NO:1, position 27645 of SEQ ID NO:1, position 32163 of SEQ ID NO:1, and position 32614 of SEQ ID NO:1.
In another embodiment, the invention provides a kit comprising at least one oligonucleotide primer pair complementary to a polymorphic region of the nucleic acid of SEQ ID NO:1, wherein the region corresponds to a polymorphic site selected from the group consisting of position 24801 of SEQ ID NO:1, position 24941 of SEQ ID NO:1, position 27645 of SEQ ID NO:1, position 32163 of SEQ ID NO:1, and position 32614 of SEQ ID NO:1.
The invention is also embodied in a method of diagnosing predisposition to left ventricular diastolic heart failure in a human, said method comprising the steps of obtaining a nucleic acid sample from the human; detecting the presence or absence of at least one allelic variant of a polymorphic region in a nucleic acid having a sequence as set forth in SEQ ID NO:1 in the sample, wherein the polymorphic region corresponds to the polymorphic site at position 24801 of SEQ ID NO:1.
The invention is also embodied in a method of diagnosing predisposition to left ventricular diastolic heart failure in a human, said method comprising the steps of obtaining a nucleic acid sample from the human; and detecting the presence or absence of at least one allelic variant of a polymorphic region in a nucleic acid having a sequence as set forth in SEQ ID NO:1 in the sample, wherein the polymorphic region corresponds to the polymorphic site at position 24941 of SEQ ID NO:1.
In another embodiment, the invention provides a method of diagnosing predisposition to left ventricular systolic heart failure in a human who has experienced a myocardial infarction, said method comprising the steps of obtaining a nucleic acid sample from the human; and detecting the presence or absence of at least one allelic variant of a polymorphic region in a nucleic acid having a sequence as set forth in SEQ ID NO:1 in the sample, wherein the polymorphic region corresponds to the polymorphic site at position 32614 of SEQ ID NO:1.
In a further embodiment, the invention provides a method of diagnosing predisposition to left ventricular diastolic heart failure in a human comprising the steps of obtaining a nucleic acid sample from the human; and detecting the presence or absence of a haplotype of the nucleic acid having a sequence as set forth in SEQ ID NO:1, said haplotype being characterized by: a G nucleotide at position 24801 of SEQ ID NO:1; a C nucleotide at position 24941 of SEQ ID NO:1; a C nucleotide at position 27645 of SEQ ID NO:1; a C nucleotide at position 32163 of SEQ ID NO:1; and a G nucleotide at position 32614 of SEQ ID NO:1.